Interaction of bleomycin, hyperthermia and a calmodulin inhibitor (trifluoperazine) in mouse tumour cells: Ii. DNA damage, repair and chromatin changes

Abstract

Summary We have reported in the preceding paper that the treatment of plateau phase mouse EMT6 tumour cells with a combination of hyperthermia (HT; 44°C) and trifluoperazine (TFP; 30, an inhibitor of calmodulin) greatly enhances the cytotoxicity of the antitumour drug belomycin (BLM). The cytotoxic action of BLM is thought to arise from the induction of DNA damage in a manner which reflects chromatin accessibility. Thus we have studied the effects of the two modifiers (HT and TFP) on chromatin structure and BLM-induced DNA damage. Co-treatment of cells with HT and TFP altered chromatin organisation by the formation and slow resolution of new DNA attachment sites at the nuclear matrix. HT increased drug-induced DNA damage (strand breaks and alkali-labile lesions) by the general depression of repair rather than through the generation of new sites for drug action. TFP produced a more discrete block in the repair of alkali-labile lesions in DNA. Both processes appear to occur for the combination of BLM, HT and TFP, and we propose that the novel chromatin configuration permits the accumulation of potentially lethal DNA strand breaks. Our study indicates the potential value of chromatin/DNA repair modifying regimens for overcoming the poor responsiveness of some tumour cells to chemotherapeutic drugs and provides a rational basis for the use of calmodulin inhibitors in thermochemotherapy.The anti-tumour glycopeptidic bleomycins are thought to exert their cytotoxic effects by damaging cellular DNA. A number of molecular mechanisms have been proposed (for review see Hecht, 1979) to explain the ability of bleomycin (BLM) to induce the liberation of free bases and the formation of single-and double-strand breaks. A central feature of recent models is the capacity of the antibiotic both to associate with DNA and to complex with ferrous iron. The ferrous oxidase activity of this complex reduces molecular oxygen to the superoxide radical, hydrogen peroxide and the potentially damaging hydroxyl radical (Caspary et al., 1982).In isolated DNA, BLM has been shown to induce single-strand breaks, alkali-labile sites (representing sites of base loss without cleavage of the phosphodiester bond) and double-strand breaks in the proportion of 5:5:1 (Lloyd et al., 1978). However, various problems are encountered in interpreting the significance of bleomycin-induced DNA damage in intact eukaryotic cells. For example, residual bleomycin may interact with DNA during the preparation of cells for damage analysis (Cox et al., 1974) and the fact that the Correspondence: P.J. Smith.

Calmodulin Mediates DNA Repair Pathways Involving H2AX in Response to Low-Dose Radiation Exposure of RAW 264.7 Macrophages

“…In this respect, prior measurements have demonstrated that CaM and histone H2AX interact to form a protein complex that dissociates upon cellular exposure to high doses of radiation , suggesting a possible functional involvement for CaM in promoting DNA repair. Alternatively, CaM may act to diminish rates of DNA damage through a variety of mechanisms that either reduce rates of reactive oxygen species generation or act to promote alterations in chromatin structure to diminish the sensitivity of DNA to oxidative damage ,, .…”

Chromosomal response of insect and mammalian cells to streptonigrin: A comparative study

“…There is abundant information indicating that the chromosomal sensitivity of mammalian cells to clastogenic compounds is closely related with the chromatin fibril structure [Kuo and Hsu, 1979;Kuo, 1981;López-Larraza and Bianchi, 1993;Smith et al, 1986]. Recently, we analyzed the sensitivity to SN of CHO chromosomes substituted with different halogenated compounds and found that DNA substitution with BrdUrd or IdUrd potentiates the chromosome damage induced by the antibiotic.…”

DNA response to bleomycin in mammalian cells with variable degrees of chromatin condensation

“…BLM induces a spectrum of DNA modifications that comprise alkali-sensitive lesions, ssDNA breaks, and dsDNA breaks in a proportion of 5:5:1 (Smith et al. 1986).…”